The TopTaq Master Mix Kit offers maximum convenience by combining all the benefits of specific TopTaq DNA Polymerase with the advantage of a ready-to-use master mix. It contains TopTaq DNA Polymerase, the unique TopTaq PCR Buffer that minimizes the requirement for optimization, and dNTPs. The unique TopTaq Stabilizer included in the enzyme storage buffer allows storage of TopTaq Master Mix at 4°C and reaction setup at room temperature, without time-consuming thawing of reagents. Providing all components in a ready-to-use master mix reduces pipetting steps, which lowers the risk of contamination. High yields of PCR product are achieved, even after storing the TopTaq Master Mix for extended periods of time at 25°C or 4°C.

The TopTaq Master Mix Kit is intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease.

Reliable high-yield PCR, independent of storage conditions.

PCR was performed using TopTaq Master Mix stored at –20°C, 4°C, and 25°C for 4 months. Two different genes from human genomic DNA (prp, 750 bp and HugL, 1200 bp) were amplified according to the standard optimized protocol (duplicates shown). The results show high yields of specific PCR product. [A] Gel image produced by the QIAxcel System. Markers: GelPilot 1 kb Plus Marker and QX Alignment Marker 15 bp/10 kb. [B] The QIAxcel System fully automates sensitive, high-resolution capillary electrophoresis of up to 96 samples per run.

Performance

TopTaq Master Mix Kit is used for standard and specialized end-point PCR applications without the need for optimization. High yields of PCR product are achieved, even after storing TopTaq Master Mix for 4 months at 25°C or 4°C, demonstrating the fact that TopTaq Master Mix offers a very robust and reliable amplification system (see figure Reliable high-yield PCR). TopTaq Stabilizer binds to TopTaq DNA Polymerase, preventing polymerase inactivation during long-term storage. With its balanced potassium and ammonium ions, the unique PCR buffer included in the master mix promotes specific primer–template annealing and simultaneously reduces nonspecific annealing. TopTaq DNA Polymerase outperformed DNA polymerases tested from other suppliers and ensures high yields of specific products, even when using low template amounts, and independent of fragment size (see figure High PCR product yields without optimization).

Principle

TopTaq Master Mix Kit has been developed by QIAGEN to provide highly reliable end-point PCR with unrivaled ease of use. Until the development of the TopTaq PCR system, all PCR enzymes required storage at –20°C. Due to the unique proprietary TopTaq Stabilizer contained in the enzyme storage buffer, TopTaq Master Mix Kit is the first PCR kit that can be stored at 4°C. This ready-to-use master mix format ensures greater convenience. High yields of specific products are achieved due to TopTaq DNA Polymerase, even when amplifying fragments of different sizes (see figure "High PCR product yields without optimization").

TopTaq PCR Buffer

The innovative TopTaq PCR Buffer facilitates the amplification of specific PCR products. During the annealing step of every PCR cycle, the buffer greatly increases the ratio of specific primer binding over nonspecific primer binding. Owing to a uniquely balanced combination of KCl and (NH4)2SO4, the PCR buffer provides stringent primer-annealing conditions over a wider range of temperatures and Mg2+ concentrations than conventional PCR buffers (see figure "Increased specificity of primer annealing"). The need to optimize PCR by varying the annealing temperature or the Mg2+ concentration is dramatically reduced, or often not required.

TopTaq Stabilizer

TopTaq Stabilizer binds to TopTaq DNA Polymerase, preventing polymerase inactivation during long-term storage (see figure "Stabilization at 4°C"). Template DNA and primers are also prevented from binding to the polymerase at low temperatures. During the initial denaturation step, TopTaq Stabilizer freely dissociates from the polymerase without compromising polymerase activity.

The TopTaq Master Mix Kit offers all of the benefits of specific TopTaq DNA Polymerase combined with the advantage of a ready-to-use master mix. With the TopTaq Master Mix Kit, separate pipetting of individual components is not required and the risk of contamination and pipetting variability is minimized, making PCR setup quick and easy. During the initial denaturation step, the TopTaq Stabilizer freely dissociates from the polymerase without compromising polymerase activity. This added convenience allows immediate reaction setup without the use of ice — saving the time usually required for reagent thawing (see flowchart "TopTaq procedure").

Reliable high-yield PCR without the need for optimization

The optimized protocol suitable for all PCR assays eliminates the need for lengthy optimization of experimental parameters for individual primer–template systems, resulting in substantial time and cost savings. High yields of specific PCR product are achieved, even after storing the TopTaq Master Mix for 4 months at 25°C or 4°C (see figure "Reliable high-yield PCR").

Applications

The TopTaq Master Mix Kit is highly suited to all end-point PCR applications.